2011 Annual Report
1a.Objectives (from AD-416)
US Veterinary Immune Reagent Network (US VIRN) www.vetimm.org was established in 2005 and represents a broad community plan to systematically address the immunological reagent gap for the U.S. veterinary immunology research community. It includes the following groups: ruminants (concentrating on cattle), swine, poultry (primarily chickens with some evaluation of reagents for turkeys), horses, and aquaculture species (concentrating on channel catfish and trout). For this renewal the groups will develop sets of reagents, i.e. monoclonal antibodies (mAb), that can identify the major leukocyte cell populations (T and B lymphocytes, macrophages, dendritic cells, neutrophils) to evaluate changes during disease and to allow scientists to manipulate them in vitro and in vivo. Such manipulations enable evaluation of the cell population’s role in protective immunity and immunopathology. In addition they will clone and expreess recombinant cytokines and chemokines (the soluble messenger molecules of the immune system) using mammalian, yeast, and bacterial systems. They will develop mAb to these proteins and their receptors to facilitate understanding of their contributions to disease and pathology. Finally, reagents to identify immunoglobulin (Ig) isotypes are needed since isotypes differ from one another functionally and thus in their effect on disease outcome. The reagents will benefit a large group of US and international researchers including veterinary immunologists, pathologists and microbiologists.
1b.Approach (from AD-416)
Prioritization of reagents (tools) to be produced will be established by surveying experts within the relevant scientific community and commercial companies as well as cooperating with similar international efforts to avoid redundancy and target the most useful reagents for research scientists. These reagents will include.
1)Cytokines: bioactive recombinant proteins as well as antibodies to them;.
2)mAbs to: Cytokine receptors, Ig isotypes, T cell receptor (TCR) constant and variable regions, Toll like (TLR) receptors, and other pattern recognition receptors and regulatory receptors including NKG2 family members and KIR. The mAb will have the following properties:.
1)For anti-cytokines mAb: mAb pairs for ELISA, ELISpot and fluorescent microsphere immunoassay (FMIA), or Luminex, assays. For these assays mAb need to react with 2 different epitopes on the same molecule. Additionally single mAb are needed for intracellular staining for cytokines;.
2)For anti-receptor mAb: mAb that block function and signaling molecules (e.g. TLR); and.
3)For cell surface or CD molecule mAbs: mAb that react with native molecules to stain viable cells for flow cytometric analyses; as well as mAb that react in fixed tissue sections. The reagents will be provided to the scientific community by depositing in commercial cell banks and encouraging generation and distribution through commercial vendors.
This was funded through a NIFA grant to the Univ. of Massachusetts. Under this renewed agreement swine immune reagents were developed to assist researchers to probe immune responses to infectious diseases and to improve vaccine design. VIRN (www.vetimm.org) was established to address the dearth of immunological reagents for food animal species. ARS Researchers at Beltsville, MD, working with Kingfisher Biotech Inc. (www.kingfisherbiotech.com/) have expressed several new recombinant swine immune proteins in yeast, including recombinant chemokines and cytokines (CCL22, CXCL9, IL-6, IFN-beta, IL-22). Bioassays have been performed at BARC to affirm that they are bioactive. Efforts are underway with our collaborators at U. Massachusetts to use these proteins for immunizations and monoclonal antibody development. Products generated by the VIRN will be used by animal health researchers, veterinarians, vaccine manufacturers, and other commercial sources. (NP103 2C)
The VIRN website for swine immunological reagents has continued to be expanded and updated. ARS Researchers at Beltsville, MD, have provided all documentation and knowhow for the USDA CSREES grant supported immune toolkit or VIRN website (http://www.umass.edu/vetimm/swine/index.html) swine pages. They scanned commercial websites and publications for swine immune reagents and organized the data into easily accessible Tables of currently available reagents for swine immunology. This list served as the basis for updating the Swine priorities for developing new VIRN immune reagents and for VIRN colleagues and swine collaborators to develop plans for their production. They also posted all current results and updated Protocols on the VIRN website. As a result this website enables researchers worldwide to quickly identify available immune tools for their experiments. Overall swine disease and vaccine researchers have improved tools to address pig health and vaccine design. (NP103 2C)
Communications for this grant have been supported by ARS by providing conference call line access for the monthly calls that review Network plans and progress. Email and phone communications are regularly exchanged between the participating labs as specific reagents are developed and screened.
Hudgens, E., Tompkins, D., Boyd, P., Lunney, J.K., Horohov, D., Baldwin, C. 2011. Expressed gene sequence of the IFN-gamma-response chemokine CXCL9 of cattle, horses, and swine. Veterinary Immunology and Immunopathology. 141(3-4):317-321.